Prostate cancer (PCa) is deadly once it metastasizes and continues to be an unmet medical need here in the United States. Increasing evidence has demonstrated that the prostate tumor microenvironment which, surrounds the cancer cells, significantly contributes to its progression, circumvention of current therapies, resistance to newer immune checkpoint therapies, and its survival. Using clinical samples, the principal investigator has found that the tumor microenvironment of metastatic castration-resistant prostate cancer (mCRPC) is infused with reactive stroma enriched with M2 macrophages known as M2-tumor-associated macrophages (M2-TAMs). His previous studies have partly elucidated how M2-TAMs and reactive stroma associate with primary and metastatic disease, identified five new surface-enriched markers on M2-TAMs, and has helped demonstrate that M2-TAMs are key regulators of PCa epithelial-to-mesenchymal transition (EMT) and tumorigenesis. Therefore, the need to design targeted therapies that target M2-TAMs within the tumor microenvironment is sorely needed. Other studies have demonstrated the importance of M2-TAMs in angiogenesis, metastasis, and their dependence on glutamine metabolism. Based on his data and that of other groups, the principal investigator hypothesizes that M2-TAMs are ideal therapeutic targets in mCRPC and help confer mCRPC resistant to immunotherapy. This proposal will determine novel M2-TAM marker expression in mCRPC (Specific Aim 1) and assess if the elimination of M2-TAMs in prostate cancer can reverse mCRPC immunotherapy resistance (Specific Aims 2, 3). This will be the first body of work using syngeneic mCRPC tumor models treated with either anti-CD206 peptide (RP182) or a novel glutamine antagonist, JHU083 followed by immunotherapy and a comprehensive immunological analysis of the tumor microenvironment. The principal investigator will also learn new techniques necessary to accomplish the proposed research under the advisement team (Drs. Drake, McConkey, Pardoll, Pienta, and Powell) and his pathologist consultant (Dr. De Marzo) all of whom have pioneering expertise in PCa biology, mCRPC treatment, cancer metabolism, single-cell RNA sequencing, and cancer immunotherapy. Importantly, his advisory committee collectively has a very strong track record of training both clinical and postdoctoral fellows who have been successful in transitioning into independent investigators at top tier research institutions. He will also engage in and present at national seminars, take coursework on laboratory biostatistics, cancer metabolism and immuno-metabolism, single cell RNA-sequencing, Jr. Faculty leadership program, grant-writing seminars, and training on running a laboratory. Combining these new skills learned during the K22 award period with his prior training in cancer biology and cancer immunology, will ensure a strong technical foundation to launch an independent laboratory dissecting and targeting innate immune cells and their mechanisms responsible for immunotherapy resistance, both of which are very poorly defined in mCRPC.